Proteomic Profiling of NAc Synaptosomes During Early and Extended Withdrawal from Self-administered Cocaine

Addiction is a devastating disorder that is exceptionally difficult to treat due to the high vulnerability of patients to relapse even long after terminating drug use. Throughout extended abstinence, cravings for drugs of abuse remain strong and even intensify, or “incubate”, over time. Craving is thought to be mediated by dysregulated signaling in the nucleus accumbens (NAc) that primes motivational circuits to initiate drug-seeking behaviour. Understanding how drug-induced synaptic changes within the NAc contribute to craving may be the key to developing effective treatments for substance abuse. The goal of the present studies is to identify protein networks that stabilize synaptic changes within the NAc following cocaine self- administration that may sensitize the system for relapse to drug-seeking behaviour. To accomplish this, we will perform cutting-edge iTRAQ discovery proteomics on synaptosomes isolated from the NAc of mice following either short- (24h) or long-term (45 days) withdrawal from cocaine self- administration. One caveat to using the iTRAQ approach is that a broad screen of the protein content of the synaptosomal fractions may not detect changes to low-abundance proteins. Therefore, an additional aim of these studies is to perform targeted analysis of protein changes within the synapse during withdrawal using the PSD/PRM approach developed by the Yale/NIDA Neuroproteomics Center. This PSD/PRM approach enables a detailed characterization of targeted protein changes within the synapse by monitoring a pre-determined set of proteins specific to the synapse. Together, these studies will identify novel changes to proteins that regulate cellular and physiological processes that can be targeted to better understand the neural mechanisms of drug craving. The results of these studies will have broad utility as a reference proteome of NAc synapses during withdrawal for future studies from our lab and others, and the use of mice provides a foundation on which to manipulate protein targets in a cell-type specific manner.